This study aimed to investigate the efficacy of gene therapy for treating autoimmune sensorineural hearing loss (ASHL) via local administration of a recombinant adenovirus vector containing the Fas ligand or interleukin IL-10 gene. Guinea pigs were divided into four groups, with different microinjections in the scala tympani. Group A were injected with FasL-EGFP, B with IL-10-EGFP, C with EGFP, and D with artificial perilymph. Seven days later, auditory brain-stem response (ABR) was tested, and the temporal bone was stained and observed by light microscopy. The spiral ligament and basement membrane were observed using transmission electron microscopy. FasL and IL-10 expression were examined using immunofluorescence histochemistry. Immunohistochemical analysis showed that the recombinant adenovirus vector in Groups A, B, and C can transfect the stria vascularis, the spiral ligament, the organ of Corti, the spiral ganglion, the region surrounding the small blood vessel in the modiolus, and the cochlear bone wall. Compared with those in Groups C and D, the ABR wave III mean thresholds were significantly lower and the inner ear immunoinflammatory responses in Groups A and B were significantly alleviated. Inhibition of immunoinflammatory response alleviated immunoinflammatory injury and auditory dysfunction. This technique shows potential as a novel therapy for ASHL.